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Abstract

The sodium channel in the heart (NaV1.5) is regulated by interactions between calmodulin (CaM), its binding motif IQ (IQ) and an upstream EF-hand like (EFL) motif in C-terminus (CT). Mutations in either IQ or EFL have been associated with long QT syndrome, suggesting the possibility of regulatory cross talk between the two motifs. Donor dequenching FRET efficiency between EYFP fused to cytoplasmic tail of NaV1.5 and ECFP conjugated with CaM or mutant CaM1234 in HEK293 cells was 2% and 5.5% respectively indicating a direct interaction and pre-association between the NaV1.5 and, CaM or CaM1234. CaM binding to NaV1.5 does not alter steady-state availability (Fig 1⇓). However truncation of CT proximal to the IQ motif, NaV1.51885, shifts the V1/2 of inactivation by −10 mV (Fig 2⇓). Mutations in the four EFL residues E1788A, D1790A, D1792A, E1799A, NaV1.54X shifts the V1/2 of inactivation −8 mV or +12 mV in the presence of CaM or CaM1234 co-expression respectively, compared to expression of NaV1.54X alone (Fig 3⇓). The effect of CaM and CaM1234 on channel inactivation is abolished by IQ/AA mutations, NaV1.54X-IQ/AA(Fig 4⇓). We conclude that the IQ motif has a critical role in tuning NaV1.5 availability through direct binding of CaM and interacting with the EFL motif in the CT.